Secretory cells are cells that are characterized by secreting biological products, such as, but not limited to, hormones (e.g., insulin), growth factors, cytokines, and so forth. Their role in biological processes is well known, and need not be set forth here. A number of diseases and pathological conditions are related to a failure of the secretory cells to work properly, such as a deficient production of the secretory products, e.g. hypothyroidism and cretin dwarfism, both due to thyroid hormone deficiency, hypophysial dwarfism due to pituitary growth hormone deficiency. Lesch-Hyhan Syndrome due to hypoxanthine-guanine phosphoribosyltransferase deficiency, fulminant hepatic failure due to the hepatotrophic factor deficiency, extracellular matrix disease due to chondrocyte deficiency, and insulin dependant diabetes due to insulin deficiency.
One approach to treat such conditions is to transplant the secretory cells into the patient. The transplanted material, in order to be clinically safe and effective, must (1) be non-immunogenic, non-thrombogenic, bio-stable, and completely non-toxic to cells and tissues of the host, (2) maintain cell viability for an extended period of time, (3) permit free passage of nutrients, secretagogues (a substance that stimulates secretion), and cell products, (4) facilitate surgical implantation and cell reseeding, and (5) be easily fixed in place and, likewise, removed.
Pancreatic islet transplantation to treat insulin-dependent diabetes has been the subject of renewed interest due to technological advances in isolating islets of Langerhans. By way of background, the human pancreas contains islets of Langerhans (hereinafter “pancreatic islets”) that are scattered throughout the exocrine pancreas with some concentrations near the pancreatic ducts. The pancreatic islets, taken together, can be thought of as a single endocrine organ occupying around 1% of the volume of the pancreas. Within the pancreas, small islets (up to 160 μm diameter) tend to distribute throughout the exocrine tissue. These small islets represent 75% of the islets in number but only about 15% in volume. Islets greater than 250 μm diameter constitute only 15% of the total number of islets but 60% of the volume. These islets are localized near larger duets and blood vessels, and are not surrounded by acinar tissue. A human pancreas may contain over 1 million islets, and each islet typically consists of several thousand cells. Each islet is comprised of a central core of insulin producing beta cells (B-cells) and a surrounding mantle of glucagon containing alpha cells (A-cells), somatostatin secreting delta cells (D-cells) and pancreatic polypeptide containing cells (PP-cells). Insulin producing B-cells makeup the majority of the cells, and comprise up to about 80% of the islets in a human.
The clinical applications of pancreatic islet transplantation have been limited to the inability to prevent islet allograft-xenograft rejection, i.e., a rejection of the transplanted pancreatic islets due to the host's immune system attacking the transplanted pancreatic islets. To counteract the rejection, the pancreatic islets have been transplanted in combination with the administration of immunosuppressive agents.
Immunosuppressive therapy, however, has proved to be a double-edged sword; while reducing the risk of rejection, it impairs the body's overall immunological defenses. Various methods of protecting the transplanted tissue from the host immune response have been explored by many investigators. As discussed below, although temporary success has been reported (See hey, Diabetes Reviews 1 (1):76 (1993), effective long-term methods have yet to be achieved.
The five major approaches to protecting the transplanted tissue from the host's immune response all involve attempts to isolate the transplanted tissue from the host's immune system. The immunoisolation techniques used to date include: extravascular diffusion chambers, intravascular diffusion chambers, intravascular ultrafiltration chambers, microencapsulation, and macroencapsulation. All of these methods have failed, however, due to one or more of the following problems; a host fibrotic response to the implant material, instability of the implant material, limited nutrient diffusion across semi-permeable membranes, secretagogue and product permeability, and diffusion lag-time across semi-permeable membrane barriers.
For example, a microencapsulation procedure for enclosing viable cells, tissues, and other labile biological membranes within a semipermeable membrane was developed by Lira in 1978. (Lim. Research report to Damon Corporation (1978)). Lim used microcapsules of alginate and poly L-lysing to encapsulate the islets of Langerhans. In 1980, the first successful in vivo application of this novel technique in diabetes research was reported ((Lim, et al., Science 210:908 (1980)). The implantation of these microencapsulated islets of Langerhans resulted in sustaining a euglycemic state in diabetic animals. Other investigators, however, repeating these experiments, found the alginate to cause a tissue reaction and were unable to reproduce Lira et al's results (Lamberti, et al., Applied Biochemistry and Biotechnology 10:101 (1984); Dupuy, et al., Jour. Biomed. Material and Res. 22:1061 (1988); Weber, et al. Transplantation 49:396 (1990); and Soon-Shiong, et al., Transplantation Proceedings 22:754 (1990)). The water solubility of these polymers is now considered to be responsible for the limited stability and biocompatibility of these microcapsules in vivo ((Dupuy, et al. supra, Weber, et al. supra, Soon-Shiong, et al., supra, and Smidsrod, Faraday Discussion of Chemical Society 57:263 (1974)).
Recently, Iwata et al., (Iwata, et al. Jour. Biomedical Material and Res. 26:967 (1992)) utilized agarose for microencapsulation of allogeneic pancreatic islets and discovered that it could be used as a medium for the preparation of microbeads. In their study, 1500-2000 islets were microencapsulated individually in 5% agarose and implanted into streptozotocin-induced diabetic mice. The graft survived for a long period of time, and the recipients maintained normoglycemia indefinitely.
Their method, however, suffers from a number of drawbacks. It is cumbersome and inaccurate. For example, many beads remain partially coated and several hundred beads of empty agarose form. Additional time is thus required to separate encapsulated islets from empty beads. Moreover, most of the implanted microbeads gather in the pelvic cavity, and a large number of islets are required in completely coated individual beads to achieve normoglycemia. Furthermore, the transplanted beads are difficult to retrieve, tend to be fragile, and will easily release islets upon slight damage.
A macroencapsulation procedure has also been tested. Macrocapsules of various different materials, such as poly-2-hydroxyethyl-methacrylate, poly vinylchloride-co-acrylic acid, and cellulose acetate were made for the immunoisolation of islets of Langerhans. (See Altman, et al., Diabetes 35:625 (1986); Altman, et al., Transplantation American Society of Artificial Internal Organs 30:382 (1984); Ronel, et al., Jour. Biomedical Material Research 17:855 (1983); Klomp, et al., Jour. Biomedical Material Research 17:865-871 (1983)). In all these studies, only a transitory normalization of glycemia was achieved.
Archer et al., Journal of Surgical Research, 28:77 (1980), used acrylic copolymer hollow fiber to temporarily prevent rejection of islet xenografts. They reported long-term survival of dispersed neonatal murine pancreatic grafts in hollow fibers which were transplanted into diabetic hamsters. Recently hey et al., Science 254:1782-1784 (1991) confirmed their results, but found the euglycemic state to be a transient phase. They found that when the islets are injected into the fiber, they aggregate within the hollow tube and result in necrosis in the central portion of the islet masses. The central necrosis precluded prolongation of the graft. To solve this problem, they used alginate to disperse the islets in the fiber. Using this method they were able to achieve long-ten graft survival. However, this experiment has not been extensively repeated. Therefore, the membrane's function as an islet transplantation medium in humans is questionable.
Thus, there exists a need for achieving secretory cell transplantation, and in particular, pancreatic islet allograft and xenograft survival without the use of chronic immunosuppressive agents.
The inventors have surprisingly discovered that macroencapsulating secretory cells in a hydrophilic gel material results in a functional, non-immunogenic, macrobead that can be transplanted into animals and can be stored for long lengths of time. The macroencapsulation of the secretory cells of the present invention provides a more effective and manageable technique for secretory cell transplantation. The macroencapsulation technique can also be used to macroencapsulate other biological agents, such as enzymes, microorganisms, trophic agents including recombinantly produced trophic agents, cytotoxic agents, and chemotherapeutic agents. The macroencapsulated biological agents can be administered to treat conditions known to respond to the biological agent.